A sepharose derivative coupled with a leupeptin-like peptide aldehyde, glycylglycyl-L-argininal, and its use as an affinity adsorbent for trypsin.
A Sepharose derivative containing a peptide aldehyde, glycylglycyl-L-argininal, the structure of which resembles that of leupeptin was prepared. It was a strong affinity adsorbent for trypsin (EC 3.4.21.4). Bovine trypsin showed higher affinity for this adsorbent at the optimum pH of catalysis (8.2) than at lower pH (5.0). This observation was in good agreement with the pH dependence of the interaction of leupeptin and trypsin (Kuramochi, H., Nakata, H. and Ishii, S. (1979) J. Biochem. 86, 1403-1410). Streptomyces griseus trypsin was also adsorbed while trypsinogen, alpha-chymotrypsin and TLCK-trypsin were not adsorbed. Though anhydrotrypsin, in which Ser-183 is converted to dehydroalanine, was not adsorbed, carbamoylmethylated (His-46) trypsin was adsorbed. Ser-183 proved to be essential for the binding. This adsorbent can also be used as a good tool to study the mechanism of action of leupeptin.
D000327:Adsorption; D002846:Chromatography, Affinity / Q000379:methods*; D006639:Histidine / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007976:Leupeptins; D009842:Oligopeptides; D012685:Sepharose / Q000031:analogs & derivatives; D012694:Serine / Q000378:metabolism; D013305:Streptomyces griseus / Q000201:enzymology; D014357:Trypsin / Q000302:isolation & purification*
D007976:Leupeptins; D009842:Oligopeptides; D012694:Serine; D006639:Histidine; C031918:glycyl-glycyl-argininal; D012685:Sepharose; D014357:Trypsin